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Intermolecular G-quadruplex induces Hyaluronic Acid-DNA superpolymers causing cancer cell swelling, blebbing and death Nathan Beals, Michael Model, Matt Worden, Torsten Hegmann, and Soumitra Basu ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.7b16983 • Publication Date (Web): 05 Feb 2018 Downloaded from http://pubs.acs.org on February 7, 2018
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ACS Applied Materials & Interfaces
Intermolecular G-quadruplex induces Hyaluronic AcidDNA superpolymers causing cancer cell swelling, blebbing and death
Nathan Beals1, Michael A Model2, Matt Worden3, Torsten Hegmann3, and Soumitra Basu1*
1
Department of Chemistry and Biochemistry, 2Department of Biological Sciences, 3Liquid
Crystals Institute, Kent State University, Kent, OH 44242
Keywords: G-quadruplex, hyaluronic acid, cancer, CD44, polymer therapeutic
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Abstract Over the past decade, nanomedicine has gained considerable traction through its relevance, for example in “smart” delivery, thus creating platforms for novel treatments. Here we report a natural polymer-DNA conjugate that undergoes self-assembly in a K+ dependent fashion to form a G-quadruplex (GQ) and generate superpolymeric structures. We derivatized a thiolated conjugate of the naturally occurring glycosaminoglycan polymer hyaluronic acid (HASH) with short G-rich DNA (HASH-DNA) that can form intermolecular non-canonical GQ structure. Gel mobility shift assay and Circular Dichroism measurements confirmed HASH conjugation to DNA and K+ dependent GQ formation respectively. TEM and SEM results indicated that the addition of K+ to the HASH-DNA conjugate led to the formation of micron range structures, while control samples remained unordered and as nebulous globular form. Confocal microscopy of a fluorescently labeled form of the superpolymer verified increased cellular uptake. The HASH-DNA conjugates showed toxicity in HeLa cells while a scrambled DNA (Mut) conjugate HASH-Mut showed no cytotoxicity, presumably due to non-formation of the superpolymeric structure. To understand the mechanism of cell death and if the superpolymeric structure is responsible for it, we monitored the cell size and observed an average of 23% increase in size compared to 4.5% in control cells at 4.5 hours. We believe that cellular stress is generated presumably by the intracellular assembly of this large superpolymeric nanostructures causing cell blebbing with no exit option. This approach provides a new strategy of cellular delivery of a targeted naturally occurring polymer and a novel way to induce superpolymeric structure formation that acts as a therapeutic.
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ACS Applied Materials & Interfaces
Introduction The CD44 receptor overexpression is found in a number of different cancer subtypes, where the receptor bound to its cognate ligand promotes the epithelial-mesenchymal transition contributing to metastasis, tumor invasion and reoccurrence 1-4. CD44 overexpression has been found to be a poor prognostic marker for prostate 5, glioma 6 and breast cancers7-10. Different isoforms of the receptor have been associated with increased tumor size, as metastatic markers in HER2/Neudependent breast cancer11 and increased tumor microenvironment heterogeneity in Triple Negative breast cancer (TNBC)7. Hyaluronic acid (HA) is an essential part of the extracellular matrix (ECM), and natural ligand for the CD44 receptor. The ligand binding to the receptor leads to cell-cell and cell-ECM interactions 12-15. The mechanism of action of HA is shown to be dependent on size, as larger high molecular weight HA fragments (HMWHA) (>106 Da) provide structural support by regulating cell adhesion, motility and growth 16. In cases of high inflammation and tumor progression, the ECM is disrupted, initiating an increase in low molecular weight HA (LMWHA) (
